The objective of this study is to develop HIV-1 vaccines targeting broadly neutralizing epitopes. Broadly neutralizing antibodies (bNAb) have been shown to be able to block infection in HIV-1 infection models as such the development of such antibodies is believed to be a vital component of any HIV-1 vaccine strategy. These bNAbs target conserved regions of HIV-1 envelope (Env), which allow for their breadth of reactivity. The elicitation of antibodies targeting these conserved regions through vaccination, however, has proven to be difficult. Currently there are no vaccine constructs that are able to effectively elicit neutralizing antibodies to these broadly neutralizing epitopes. The first aim of this projec is to develop E2- based vaccines targeting the broadly neutralizing epitopes of the HIV-1 envelope. To complete this aim the Env regions of MPER and V1V2 will be computationally modeled onto the surface of the E2 scaffold protein. This will be done to ensure that the proper conformation is presented on the surface of the vaccine particle. Once the modeling has been completed, wild-type SF162 MPER and V1V2 as well as E2 will be modified as required to match the model antigens. Completed Env-E2 constructs will be produced and purified from mammalian cells. The constructs will be tested by Western Blot, Native PAGE, competition assay, as well as by immunoprecipitation to ensure that the V1V2 and MPER regions are presented properly on the purified Env- E2 particles. Aim 2 of this project is to determine the immunogenicity and breadth of neutralizing antibodies generated following vaccination with Env-E2 constructs. To test this aim rabbits will be vaccinated with the Env-E2 vaccines delivering a total of 4 vaccinations given at 6-week intervals. The elicitation of antibody responses will be measured by ELISA to gp120, gp41, gp140, and the Env peptide set. Additionally neutralization will be measured by TZM-bl neutralization assay to SF162, a panel of tier 1 and 2 pseudoviruses as well as HIV-2/HIV-1 (MPER and V1V2) chimera viruses. The overall evaluation of the elicited immune responses will determine the strength of antibody response generated by the vaccines as well as to determine the region(s) of Env targeted by these responses. If this project is successful it will greatly advance the field of HIV-1 vaccine development by providing a neutralizing antibody component to an overall HIV-1 vaccine strategy.